[Rapid characterization and identification of non-volatile constituents in Pogostemonis Herba by UPLC-Q-TOF-MS combined with UNIFI and an in-house library].

This study aimed to characterize and identify the non-volatile components in Pogostemonis Herba by using ultra-perfor-mance liquid chromatography-quadrupole-time of flight-mass spectrometry(UPLC-Q-TOF-MS) combined with UNIFI and an in-house library. The chemical components in 50% methanol extract of Pogostemonis Herba were detected by UPLC-Q-TOF-MS in both positive and negative MS~E continuum modes. Then, the MS data were processed in UNIFI combined with an in-house library to automatically characterize the metabolites. Based on the multiple adduct ions, exact mass, diagnostic fragment ions, and peak intensity of compounds and the fragmentation pathways and retention behaviors of reference substances, the structures identified by UNIFI were further verified and those of the unidentified compounds were tentatively elucidated. A total of 120 compound structures were identified or tentatively identified, including flavonoids, phenylpropanoids, phenolic acids, terpenes, fatty acids, alkaloids, and phenylethanoid glycosides. Sixteen of them were accurately identified by comparison with reference substances, and 53 compounds were reported the first time for Pogostemonis Herba. This study systematically characterized and identified the non-volatile compounds in Pogostemonis Herba for the first time. The findings provide a scientific basis for revealing the pharmacodynamic material basis, establishing a quality control system, and developing products of Pogostemonis Herba.

[Research progress on chemical constituents and bioactivities of Lepidium meyenii].

Maca( Lepidium meyenii) known as the " national treasure of Peru" and " South American ginseng",is annual or biennial herbs of the genus Lepidium in Cruciferae. It mainly contains proteins,amino acids,polysaccharides,alkaloids( including:macamides,imidazoles,hydroxypyridines,carbazoles,organic amines and so on),glucosinolates,macaenes,thioethylurea,sterols and other chemical constituents. In recent years,more and more studies have found that it could treat osteoporosis and improve prostatehyperplasia,and possessed anti-cancer,female climacteric syndrome,rheumatism,antioxidant and other pharmacological effects. In this paper,the chemical constituents and bioactivity of Maca were reviewed,which could provide the basis for the further development and utilization of Maca.

[Comparison of volatile components in Schisandra propinqua from different habitats by HS-SPME-GC-MS].

A headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) method was developed and optimized for qualitative and semi quantitative analysis of volatile components from Schisandra propinqua, a kind of Yi Nationality herb medicine. This method was used for analysis and evaluation of volatile components from S.propinqua from four different geometrical origins (Yunnan Wuding, Yunnan Luoping, Guizhou Qingzhen and Hubei Shennongjia). 51, 53, 52， 50 compounds were identified from the above four kinds of samples respectively, in which 46 volatile components were contained in all of these four origins. The volatile components in these samples from different geographical origins were then compared by using principal component analysis, cluster analysis and other chemometrics methods. The results suggested that the analysis of volatile components can be used to distinguish the S.propinqua from four origins, and those samples from close geographical origins were more similar in volatile components. The developed method was stable, reliable, and suitable for rapid analysis of volatile components from S.propinqua, providing reference for quality control, drug development and scientific utilization of the herb.

[Rapidly identify oligosaccharides in Morinda officinalis by UPLC-Q-TOF-MSE].

In this paper, an approach was applied for separation and identification of oligosaccharides in Morinda officinalis How by Ultra performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) with collision energy. The separation was carried out on an ACQUITY UPLC BEH Amide C18(2.1mm×100 mm，1.7 µm) with gradient elution using acetonitrile(A) and water(B) containing 0.1% ammonia as mobile phase at a flow rate of 0.2 mL·min⁻¹. The column temperature was maintained at 40 °C. The information of accurate mass and characteristic fragment ion were acquired by MSE in ESI negative mode in low and high collision energy. The chemical structures and formula of oligosaccharides were obtained and identified by the software of UNIFI and Masslynx 4.1 based on the accurate mass, fragment ions, neutral losses, mass error, reference substance, isotope information, the intensity of fragments, and retention time. A total of 19 inulin oligosaccharide structures were identified including D(+)-sucrose, 1-kestose, nystose, 1F-fructofuranosyl nystose and other inulin oligosaccharides (DP 5-18). This research provided important information about the inulin oligosaccharides in M. officinalis. The results would provide scientific basis for innovative utilization of M. officinalis.

[Rapidly identify chemical constituents of Schisandra propinqua by UPLC-Q-Tof/MSE].

This study was to identify the chemical constituents of Schisandra propinqua, one herbal medicine of Yi nationality in China by using ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF/MSE). Acetonitrile- water containing 0.1% formic acid was used as the mobile phase for gradient elution. Data were collected under ESI negative mode and ESI positive mode, and then screened and verified by the software of UNIFI and Masslynx4.1. Based on the accurate mass, fragment ions, neutral losses, mass error, retention time, reference substance, isotope information, the intensity of fragments, as well as the previous reports, the known compounds were validated and identified. The chemical structures of the unknown components were identified according to exact molecular weight, MS fragment, chromatographic retention behavior, and characteristic fragments of known congener compounds. A total of 68 chemical components were identified from S. propinqua, including 3 flavonoids, 10 flavanols, 34 lignans compounds (including 20 dibenzocyclooctene lignans), 4 triterpenoids, 17 organic acid and other compounds. 37 compounds of them were found in S. propinqua for the first time, and one potential compound needed to be identified.

[Study on distribution of five heavy metal elements in different parts of Cordyceps sinensis by microwave digestion ICP-MS].

The contents of five heavy metals (Cu, Pb, As, Cd, Hg) in 17 batches of Cordyceps sinensis were determined by microwave digestion-ICP-MS, and their distribution in C. sinensis were analyzed. The results showed that the contents of Cu, Pb, Cd and Hg in all batches were in accordance with the international standards of Chinese Medicine-Chinese Herbal Medicine Heavy Metal Limit, with their contents in the stroma higher than that in the caterpillar body, and the excess rate of As, which mainly concentrated in the caterpillar body part of C. sinensis, was 88.24%, as the content of As in the caterpillar body was 7 to 12 fold of that in the stroma. In this study, the distribution of five heavy metals in C. sinensis was clarified, and the existing problems of arsenic limit of heavy metal in C. sinensis were analyzed, and some suggestions were put forward. It is hoped that the reference standard can be provided for the limited standard of arsenic in C. sinensis.

[Climate change impacts on yield of Cordyceps sinensis and research on yield prediction model of C. sinensis].

Cordyceps sinensis is a Chinese unique precious herbal material, its genuine producing areas covering Naqu, Changdu in Qinghai Tibet Plateau, Yushu in Qinghai province and other regions. In recent 10 years, C. sinensis resources is decreasing as a result of the blindly and excessively perennial dug. How to rationally protect, develop and utilize of the valuable resources of C. sinensis has been referred to an important field of research on C. sinensis. The ecological environment and climate change trend of Qinghai Tibet plateau happens prior to other regions, which means that the distribution and evolution of C. sinensis are more obvious and intense than those of the other populations. Based on RS (remote sensing)/GIS(geographic information system) technology, this paper utilized the relationship between the snowline elevation, the average temperature, precipitation and sunshine hours in harvest period (April and may) of C. sinensis and the actual production of C. sinensis to establish a weighted geometric mean model. The model's prediction accuracy can reach 82.16% at least in forecasting C. sinensis year yield in Naqu area in every early June. This study can provide basic datum and information for supporting the C. sinensis industry healthful, sustainable development.

[Qualitative and quantitative analyses of primary saponins in Paris forrestii].

In order to study whether Paris forrestii could be developed as a substitute of Paridis Rhizome, chemical compositions of P. forrestii and P. polyphylla var. yunnanensis were investigated by UPLC-Q-TOF MS. In addition, the contents of eight primary steroidal saponins in 77 batches of P. forrestii samples from different habitats were simultaneously determined by HPLC-UV. The results showed that P. forrestii and P. polyphylla var. yunnanensis have similar chemical compositions, and all 22 major common peaks were identified as steroid derivatives. Meanwhile, there were some differences in the contents of saponins in P. forrestii samples from different habitats. The contents of 4 steroidal saponins in Chinese Pharmacopoeia ranged from 0.068% to 3.30%, and the highest content of the 8 kinds of steroidal saponins was 6.18%, while the lowest was just 0.71%. Moreover, 78% of P. forrestii samples were in conformity with the requirements of Chinese Pharmacopoeia, indicating that P. forrestii samples had relatively stable quality and could be further studied as a substitute for Paridis Rhizome.

[Determination of eight steroidal saponins in 15 kinds of genus Paris].

The study was established an HPLC method for simultaneous determination of 8 steroidal saponins (polyphyllins Ⅶ, H, Ⅵ, Ⅱ, Ⅰ, and Ⅴ, dioscin, and gracillin) in Paridis Rhizoma, and made an evaluation by determining steroidal saponins in 15 kinds of genus Paris. The analysis was performed on a Waters Acquity H-ClassTM UPLC ultrafine liquid chromatography system coupled with a PDA detector. The chromatographic separation was achieved through a CAPCELL PAK ADME (4.6 mm× 250 mm, 5 µm) column and the optimal mobile phase consisted of acetonitrile and water. The column was maintained at 21 ℃, and the flow rate was 0.8 mL•min ⁻¹. The UV detection wavelength was 203 nm. The results showed that ① the detected components can be well separated and all with good correlation coefficients. The standard calibration curves were linearly good (R2>0.999 9). The linearity was obtained over 0.041 70-3.812 00 µg. The average recoveries ranged from 95.91% to 103.8%. ② there are significant differences in the content of steroidal saponins from different species. The steroidal saponins are low content or almost none in P. mairei, P. polyphylla var. stenophylla, and P. delavayi have low content or almost did not contain, so these species are not suitable for medicinal use. The contents of steroidal saponins in P. polyphylla var. chinensis are varied from different places. There were high content of steroidal saponins in P. polyphylla var. yunnanensis, P. forrestii, P. daliensis, and P. axialis, even up to 5.0%, which indicated that they had the potential pharmic value of development.

[Advance in studying early bolting of Umbelliferae medicinal plant].

The early bolting has become one of the key issues affecting the Umbelliferae herbs production. The early bolting does not only reduce herbs production but also affect the active ingredients of plants. The concept of early bolting, physiological theory, induce factors of the quality of medicine and, control technology about the early bolting of Umbelliferae are integrative reviewed in the paper. Its occurrence mechanism include: genetic factors, environmental and ecological factors, endogenous hormones, sowing, fertilizer and other production management measures. For the above occurring mechanism, the proposed prevention measures are as follows: preferably germplasm, sowing, rational fertilization, exogenous hormones, shading, reed head cut off, cut fall the leaves, reasonable planting density and other agronomic measures can reduce its bolting rate. And the early bolting problem focused for future research are mentioned, including: development of authentic ingredients cultivation technical specifications, screened early bolting hormone inhibitor, in-depth study of the mechanism of early bolting, reveal its early bolting gene and key ecological factors. The research of vernalization gene, gene regulation of flowering early bolting will solve the problem of quality and production reducing fundamentally caused by the early bolting.

[Determination of five effective components in medicinal material of Tripterygium by UPLC-ESI-MS/MS].

This paper is aimed to develop a method for the determination of five effective components in medicinal material of Tripterygium using ultra performance liquid chromalography coupled with electrospray ionization tandem mass spectrometry(UPLC-ESI-MS/MS), which then was used to study their contents in raw materials from different areas and different sources.The separation was performed on a Waters ACQUITY UPLC-CSH-C18S column(2.1 mm×100 mm,1.7 µm), usingacetonitrile-0.2% ammonium form ateaqueous solutionas mobile phase. The target components were detected in multiple-reaction monitoring(MRM) mode by mass spectrometry with electrospray ionization (ESI) source operated in positive ionization mode. The quantitative results showed that good linearity was achieved in their respective linear ranges and fine determination coefficient (r > 0.997 8),and the overall recoveries ranged from 96.72%-103.2% with the RSD ranging from 1.0%-2.4%.The method is sensitive and accurate, and suitable for the effective components quantification in medicinal material of Tripterygium; contents of five effective components from different sources vary significantly, so the quality and safety of medicinal material of Tripterygium needs to be improved. It is very important to control the quality with multi-index for clinic safety.

[Different metabolites of leaves between Tripterygium wilfordii and Tripterygium hypoglaucum based on UPLC-Q-TOF-MS].

To analysis the differences between Tripterygium wilfordii and T. hypoglaucum, specimens of their leaves were collected from five production regions and analyzed by ultra performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-Q-TOF-MS). The data were analyzed by multivariate statistical method, such as hierarchical cluster analysis (HCA) principal component analysis (PCA) and orthogonal signal correction partial least square discrimination (OPLS-DA). Potential markers with VIP values above 5.0 and corresponding r values above 0.85, were selected and further tested by combining mann-Whitney nonparametric. Those with P < 0.001 and AUC = 1 were confirmed as metabolite markers to discriminate them from each other. Results revealed that the two species were obviously different in their leaf metabolites. Based on their mass spectra, 23 potential metabolite markers were identified to distinguish T. wilfordii from T. hypoglaucum.

[Investigation Report of Tripterygium wilfordii and Tripterygium hypoglaucum].

OBJECTIVE: To get the information of resources, cultivation, commodity circulation and other aspects of Tripterygium wilfordii and Tripterygium hypoglaucum. METHODS: Collect samples in 13 locations of Tripterygium wilfordii and Tripterygium hypoglaucum, compare their plant morphological characteristics and growth habit, and investigate their wild resources conditions, planting information, easy-confused varieties and different commodity features. RESULTS: (1) Tripterygium wilfordii and Tripterygium hypoglaucum were mainly collected under woods or on the edge of woods,and light and moisture attributed to their distribution to some extent. (2) Wild resources of Tripterygium wilfordii and Tripterygium hypoglaucum were shrinking, and both of their cultivation history were relatively short and their cultivation technique were still in a low level. (3) Due to lack of harvesting and processing standards, decoction pieces, varying from roots, rhizomes to stems of plants, were all sold as commercial medicines. CONCLUSION: Wild resources of Tripterygium wilfordii and Tripterygium hypoglaucum are shrinking,and the standardized research on cultivation-harvest processing and commercial medicines remains to be further carried out.

[Protection and bidirectional effect of rhubarb anthraquinone and tannins for rats' liver].

OBJECTIVE: To compare the bidirectional effect of rhubarb total anthraquinone (TA) and total tannins (TT) on rats' liver. METHODS: One hundred rats were randomly divided into 10 groups, i.e., the blank group, the model group, the blank + high dose TA group, the blank +low dose TA group, the blank + high dose TT group, the blank + low dose TT group, the model + high dose TA group, the model + low dose TA group, the model +high dose TT group, and the model + low dose TT group, 10 in each group. The carbon tetrachloride (CCI4) was used to prepare the acute liver injury rat model. TA and TT of rhubarb (at 5.40 g crude drugs/kg and 14.69 g crude drugs/kg) were intragastrically administrated to rats in all groups except the blank group and the model group, once daily for 6 successive days.The general state of rats, biochemical indices such as alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), laminin (LN), hyaluronic acid (HA), transforming growth factor beta1 (TGF-beta1), as well pathological results of rat liver tissues. Finally the protection laws of TA and TT for rats' liver were analyzed using factor analysis. RESULTS: Compared with the blank control group, all biochemical indices increased in the blank group (P < 0.05, P < 0.01). HA also increased in the blank + high dose TA group; AST, ALT, and HA also increased in the blank +high dose TT group (P < 0.05). Compared with the model group, AST, ALT, ALP, HA, and TGF-beta1 significantly decreased in the model + low dose TA group, the model + high dose TA group, the model + low dose TT group (P < 0.05, P < 0.01). Serum AST, ALT, and ALP also decreased in the model + high dose TT group (P < 0.05, P < 0.01). Pathological results showed that mild swollen liver cells in the model + high dose TA group. Fatty degeneration and fragmental necrosis around the central veins occurred in the blank + high dose TA group. The pathological injury was inproved in the model +low dose TA group. Two common factors, liver fibrosis and liver cell injury, were extracted by using factor analysis. TA showed stronger improvement of the two common factors than TT. CONCLUSIONS: Rhubarb TA and TT showed protective and harmful effects on rats' liver. At an equivalent dosage, TA had better liver protection than TT. High dose TT played a role in liver injury to some extent.

[Comparison of agronomic traits of Panax notoginseng between traditional cultivated fields and new cultivated fields].

OBJECTIVE: In order to provide scientific basis for introduction breeding and production regionalization of Panax notoginseng, the environment of producing area, agronomic traits and medicinal material output were investigated. METHOD: Using field survey sampling at harvest time, agronomic traits indicators of leaf segment, stem segment and roots segment etc. of commodity P. notoginseng, longitude and latitude, elevation, soil type, landform of producing area were measured and observed. RESULT: The P. notogiseng cultivation was expanding from traditional area like Wenshang to new areas as Honghe, Kunming, Qujing at large scale. Comparing with traditional cultivated fields, the elevation of new fields, which are red soil of moderate or low mountain slopes and gentle hills between 1 800-2 130 m, increases markedly. The agronomic traits of new cultivated fields such as plant height, stem diameter, the ground and underground biomass were better than those of traditional cultivated fields in varying degree. Furthermore, the root weight, taproot weight, taproot length and other agronomic index augmented more than 20%. Comparison among different cultivated fields showed P. notoginseng of Honghe Shiping Niujie, Kunming Shilin Guishan, Honghe Jiangshui Guanting etc. had better agronomic traits, the plant were thick and tall, the taproot and the top of reed were large, the number of root was more. Inversely, P. notoginseng of Kunming Guandu Xiaoshao, Guangxi Jingxi Lutong, Wenshang Yanshan Jiangna were thin and small. Cluster analysis showed that cultivated fields of P. notoginseng which was across clustered by traditional and new cultivated fields can be divided into three groups, including a high-yielding region insist of three new cultivated fields Honghe Shiping Niujie, Kunming Shilin Guishan, Honghe Jiangshui Guanting and one traditional cultivated field Wenshan Yanshan Jiangna. Correlation analysis indicated that the size and weight of taproot and top of reed were significantly positive correlated with plant height, stem diameter, leaf size, leaf weight and stem weight. Regression analysis pointed out that stem diameter, leaf width, leaf length/width, leaf weight were the main factors affecting the dry weight of taproot, and the main factors influencing the dry weight of top of reed were plant height, petiole, leaf length, leaf width and other agronomic traits. CONCLUSION: From agronomic traits, P. notoginseng in new cultivated fields were more robust than that in traditional fields, but besides the length of taproot, the difference of rest agronomic traits didn't reach the significant level. It is suggested that, in the west and the north of traditional area, the red soil land of moderate or low mountain slopes and gentle hills where the elevation is between 1 800-2 130 m is suitable for cultivating P. notoginseng. Honghe Shiping Niujie, Honghe Jianshui Guanting, Kuming Shilin Gongshan can be used as vital development area for commodity P. notoginseng.

[Determination and analysis of heavy metals content in Panax notoginseng of different origination].

Six heavy metals, including As, Cu, Hg, Cd, Pb and Cr in Panax notoginseng were determined by inductively coupled plasma atomic emission spectrometry (ICP-AES) combined with wet digestion method. The samples of P. notoginseng were collected in 12 different regions, including Yunnan and Guangxi Province. Green Standards of Foreign Trading Medicinal Plants & Preparations was used as the standard to evaluate the pollution status of As, Cu, Hg, Cd, Pb and Cr in P. notoginseng. The results showed that content of As and Cd exceeded the limit of the standard and the percentage was 32.4% and 29.7%, respectively, while Cu, Hg and Pb were all bellow the limit. The SPSS 16.0 software was used to analyze the data. The occurrence of contained heavy metals has been discussed.

Untargeted metabolite analysis-based UHPLC-Q-TOF-MS reveals significant enrichment of p-hydroxybenzyl dimers of citric acids in fresh beige-scape Gastrodia elata (Wutianma).

In order to comprehensively elucidate the chemical biosynthesis process of the beige-scape Gastrodia elata Blume (Wutianma) as a traditional herbal medicines, the untargeted analysis-based UHPLC-PDA-ESI-Q-TOF-MS reveals the metabolites ranging from the skeletons to novel dimers of citric acids in fresh and dried immature/mature stem tubers. Interestingly, two novel types of dimers for citric acids with the anhydride groups at sn-1 and/or sn-5 were discovered in fresh samples. Moreover, the classical mono- versus novel di-mers, and the aglycons versus the glycosides could be easily discriminated by signature fragmentation patterns and some novel adduct ions. The heat map of contents demonstrated more p-hydroxybenzyl metabolites than gastroxyl ones were determined in fresh Wutianma revealing a significant specificity with the lack of the sufficient gastrodin and gastroxyl products in biosynthetic pathway.

Structural characterization and discrimination of the Paris polyphylla var. yunnanensis and Paris vietnamensis based on metabolite profiling analysis.

This study aimed to distinguish the rhizomes of Paris polyphylla var. yunnanensis (Franch) Hand Mazz (PPY) and Paris veitnamensis (Takht.) H. Li (PV) using metabolomics-based ultra high-performance liquid chromatography coupled with quadrupole time-of-fligh mass spectrometry (UHPLC/Q-TOF MS). First, the UHPLC/Q-TOF MS approach was optimized for metabolite profiling. Then, the MS data were processed using UNIFI™ combined with an in-house library to automatically characterize the metabolites. Based on the exact mass information, the fragmentation characteristics, and the retention time of compounds, and the fragmentation mechanism and retention behavior of steroidal glycosides in the references, the structures identified by UNIFI were further verified. Overall, 146 metabolites, including 42 potential new compounds, were identified or tentatively identified. Pattern recognition analysis of the PPY and PV MS data revealed that they were clearly separated, and 15 potential biomarkers for differentiating between them were selected. These biomarkers were subsequently used to successfully predict the genus of PPY and PV samples. These results indicated that metabolite profiling by UHPLC/Q-TOF MS is an effective, robust approach for determining the characteristic biomarkers that differentiate between TCM species with multiple botanical origins.